There is a fundamental gap in understanding how Granulin (GRN) regulates myeloid cell differentiation. Continued existence of this gap represents an important problem because, until it is filled, understanding of how Granulin contributes to the development of acute myeloid leukemia would be unknown, and therefore the manipulation of the Granulin pathway to treat these hematopoietic malignances will remain unreachable. The long-term goal is to improve the prognosis of patients suffering from acute myeloid leukemia by expanding their therapeutic options. The overall objective is to define in vivo the myeloid cell populations that require Granulin for proper development as well as the molecular pathway activated by Granulin in myeloid cells. The central hypothesis is that Granulin is essential for proper myeloid lineage differentiation of granulocytes, neutrophils and macrophages through the activation of signal transducers and activators of transcription (STAT) family members. This hypothesis has been formulated on the basis of preliminary data produced by the applicant. The rationale for the proposed research is that understanding the fundamental molecular mechanisms of hematopoietic regulation by Granulin has the potential to translate into better understanding of the pathogenesis of acute myeloid leukemia, blood malignancies with a low cure rate of 24%. Guided by strong preliminary data, this hypothesis will be tested by pursuing two specific aims: 1) Identify in vivo the myeloid cell populations that require Grna for proper development; and 2) Determine which STAT family members are activated through Granulin for proper myeloid differentiation. Under the first aim, a published grna mutant which has been described by the applicant to have decreased myeloid cell numbers, will be used to identify in vivo the myeloid cell populations whose development is affected by the absence of Grna, both during embryonic and adult hematopoiesis. Powerful in vivo microscopy tools in the zebrafish embryo, histological techniques, and RNA-hybridization probes that are already on hand will be used. Under the second aim, quantitative PCR to pinpoint the STAT family members activated by Grna, and injection of mRNA for each STAT candidate in grna mutant embryos will be performed to rescue myeloid defects. An innovative approach is proposed by taking advantage of the zebrafish genome duplication that resulted in two copies of the ancestral Granulin gene (grna and grnb) to understand in an unprecedented manner the hematopoietic function of Granulin due to the specialization of grna in hematopoietic processes. Since grna is specific to hematopoietic processes, this avoids disruption of other tissues. In addition, the power of the zebrafish for visualization of in vivo development of hematopoiesis is exploited. The proposed research is significant, since it is expected to vertically advance understanding of how Granulin could be contributing to acute myeloid leukemia and how this protein could be manipulated to treat these disorders.